Transcriptional response to aminoglycoside antibiotics in cochlear hair cells at 3 hour
Ontology highlight
ABSTRACT: We purified Atoh1-GFP positive hair cells from organotypic cultures of P1 cochlea 3 hours after 0.5mM gentamicin treatment and performed RNA sequencing to profile the early transcriptional response of hair cells to aminoglycoside antibiotics. Levels of mRNA in gentamicin-treated hair cells (three replicates) were compared to untreated hair cells (three replicates). GFP negative, non-hair cells populations from treated organs were compared to those from untreated organs (three replicates for each condition).
Project description:We purified Atoh1-GFP positive hair cells from organotypic cultures of P1 cochlea 3 hours after 0.5mM gentamicin treatment and performed RNA sequencing to profile the early transcriptional response of hair cells to aminoglycoside antibiotics.
Project description:To investigate alterations in gene expression after Ad2-ATOH1-GFP transduction of human supporting cells, we measured transcriptome changes by RNA-seq. We compared gentamicin-treated tissue (to kill the hair cells) transduced with Ad2-ATOH1-GFP (to promote hair cell production) cultured for 5 days, versus control tissues only treated with gentamicin (cultured for 2 days, 8 days and 14-18 days).
Project description:We purified seven different cell populations and performed RNA sequencing to profile transcriptional similarities and differences between them. The seven cell types were 1) Atoh1-GFP positive cochlear hair cells from the organ of Corti of postnatal day one mice, 2) Atoh1-GFP negative cells from the organ of Corti of postnatal day one mice, 3) Atoh1-GFP positive induced hair cells generated by overexpression of Six1, Atoh1, Pou4f3, and Gfi1, 4) dsRed transduced control mouse embryonic fibroblasts, 5) Atoh1-GFP positive Merkel cells from postnatal day 1 mice, 6) Atoh1-GFP positive Cerebellar granule precursor cells from postnatal day 1 mice, and 7) Atoh1-GFP positive Gut secretory cells from postnatal day one mice.
Project description:We purified seven different cell populations and performed RNA sequencing to profile transcriptional similarities and differences between them. The seven cell types were 1) Atoh1-GFP positive cochlear hair cells from the organ of Corti of postnatal day one mice, 2) Atoh1-GFP negative cells from the organ of Corti of postnatal day one mice, 3) Atoh1-GFP positive induced hair cells generated by overexpression of Six1, Atoh1, Pou4f3, and Gfi1, 4) dsRed transduced control mouse embryonic fibroblasts, 5) Atoh1-GFP positive Merkel cells from postnatal day 1 mice, 6) Atoh1-GFP positive Cerebellar granule precursor cells from postnatal day 1 mice, and 7) Atoh1-GFP positive Gut secretory cells from postnatal day one mice.
Project description:We purified seven different cell populations and performed RNA sequencing to profile transcriptional similarities and differences between them. The seven cell types were 1) Atoh1-GFP positive cochlear hair cells from the organ of Corti of postnatal day one mice, 2) Atoh1-GFP negative cells from the organ of Corti of postnatal day one mice, 3) Atoh1-GFP positive induced hair cells generated by overexpression of Six1, Atoh1, Pou4f3, and Gfi1, 4) dsRed transduced control mouse embryonic fibroblasts, 5) Atoh1-GFP positive Merkel cells from postnatal day 1 mice, 6) Atoh1-GFP positive Cerebellar granule precursor cells from postnatal day 1 mice, and 7) Atoh1-GFP positive Gut secretory cells from postnatal day one mice.
Project description:This study examined transcripts that are enriched in neonatal mouse cochlear hair cells. Hair cells were purified by FACS sorting for GFP fluorescence from the cochleas of transgenic mice in which the endogenous Atoh1 gene was fused with GFP Two replicates of GFP+ hair cells were compared with all other cochlear cell types that were GFP-
Project description:This study examined transcripts that are enriched in neonatal mouse cochlear hair cells. Hair cells were purified by FACS sorting for GFP fluorescence from the cochleas of transgenic mice in which the endogenous Atoh1 gene was fused with GFP
Project description:The mammalian cochlea loses its ability to regenerate new hair cells prior to the onset of hearing. In contrast, the adult vestibular system can produce new hair cells in response to damage, or by reprogramming of supporting cells with the hair cell transcription factor Atoh1. We used RNA-seq and ATAC-seq to probe the transcriptional and epigenetic responses of utricle supporting cells to damage and Atoh1 transduction. We show that the improved regenerative response of the utricle correlates with a more accessible chromatin structure in utricle supporting cells compared to their cochlear counterparts. We also provide evidence that Atoh1 transduction of supporting cells is able to promote increased transcriptional accessibility of some hair cell genes. Our study offers a possible explanation for regenerative differences between sensory organs of the inner ear, but shows that additional factors to Atoh1 may be required for optimal reprogramming of hair cell fate.
Project description:Neural basic helix-loop-helix (bHLH) transcription factors are important for the differentiation and cell type specification of neurons. They are thought to share direct downstream targets in their common role as neuronal differentiation factors, but have distinct targets with respect to their cell type specific roles. Little is known about distinct cell-type specific bHLH targets as previous work did not distinguish these from common targets. Based on previous genetic evidence, we hypothesize that bHLH transcription factors have unique targets for their function in regulating neuronal sub-type specification. Atoh1 (Math1) is a bHLH transcription factor that specifies different cell types of the proprioceptive pathway in mammals such as the dorsal interneuron 1 population of the developing neural tube. Using microarray analyses of neighboring specific bHLH sorted populations from developing mouse neural tubes, we determine transcripts unique to the Atoh1-derived population and not those common to bHLH transcription factors in related neural progenitor populations. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) experiments of native tissue followed by enhancer reporter analyses identified five direct cell-type specific targets of Atoh1 in vivo: Klf7, Rab15, Rassf4, Selm, and Smad7, along with their Atoh1-responsive enhancers. These Atoh1 targets were found from native tissue in the appropriate developmental context and have diverse functions that range from transcription factors to regulators of endocytosis and signaling pathways. Only Rab15 and Selm are expressed across several different Atoh1-specified cell types including external granule cells (EGL) in the developing cerebellum, hair cells of the inner ear, and Merkel cells, demonstrating that even within Atoh1 lineages, not all Atoh1 specific targets are shared. Our work establishes on a molecular level that the neuronal differentiation bHLH transcription factors also have distinct targets for their roles in neuronal sub-type specification. From this work, we can begin to address how bHLH transcription factors are able to specify unique cell types and initiate programs that organize neuronal diversity. Gene expression analysis: Two samples, Atoh1-GFP and dNeurog1-GFP, were analyzed. Two biological replicates of each. Atoh1-GFP transgenics are mice with GFP inserted into a BAC that drives GFP to the Atoh1-derived population of the developing neural tube marking the dorsal interneuron 1 population (Raft et al. Development 2007). dNeurog1-GFP transgenics are mice with a Neurog1 enhancer that drives GFP to the dorsal part of the developing neural tube marking the dorsal interneuron 2 population (Nakada et al. Dev Bio 2004). Chip-seq analysis: Series GSE22111
Project description:Vestibular hair cells are mechanoreceptors critical for detecting head position and motion. In mammals, hair cell loss causes vestibular dysfunction as spontaneous regeneration is nearly absent. Constitutive expression of exogenous ATOH1, a hair cell transcription factor, increases regeneration of hair cells, but these cells fail to mature. With the aim of developing an optimized AAV-based ATOH1 gene therapy, we used single-cell RNA-Seq and explored the impact of defined promoters on ATOH1 transgene expression level, timing, effects in distinct cell types, and the maturity of regenerated vestibular hair cells ex vivo and in vivo.